Molecular confirmation & characterization of Rickettsia conorii in north India: A report of three cases.

Background & objectives: In India, spotted fever group rickettsiae (SFGR) are an underdiagnosed cause of acute febrile illness (AFI). The non-specific Weil-Felix test is the first diagnostic modality for the diagnosis of SFGR in many laboratories due to the lack of advanced diagnostic facilities in developing countries. The aim of this study was to detect SFGR using molecular methods in the patients, presenting with AFI in a tertiary care centre in north India. Methods: Consecutive patients (>14 yr of age) with AFI were enrolled over a six month period. Standard investigations for common pathogens causing AFI in India (malaria, dengue, scrub typhus, leptospirosis and enteric fever) were carried out. In patients who were negative for all of the above investigations, blood was subjected to polymerase chain reaction (PCR) targeting outer membrane protein A (ompA) gene of Rickettsia. Results: Of the 51 patients with an undiagnosed aetiology, three were positive by ompA PCR. Two of the PCR products produced good sequences and BLAST identification confirmed them as Rickettsia conorii. The sequences of R. conorii reported from south India clustered with two previously reported novel rickettsial genotypes. The study sequences clustered in a group different from that of Rickettsia spp. of the south Indian sequences reported earlier. Interpretation & conclusions: This study showed the existence of R. conorii in north India. Testing for SFGR may be included in the diagnostic workup of AFI for better disease management.

Rickettsia conorii O antigen is the target of bactericidal Weil-Felix antibodies.

Rickettsial diseases have long been diagnosed with serum antibodies cross-reactive against Proteus vulgaris (Weil-Felix reaction). Although Weil-Felix antibodies are associated with the development of immunity, their rickettsial target and contribution to disease pathogenesis are not established. Here, we developed a transposon for insertional mutagenesis of Rickettsia conorii, isolating variants defective for replication in cultured cells and in spotted fever pathogenesis. Mutations in the polysaccharide synthesis operon (pso) abolish lipopolysaccharide O-antigen synthesis and Weil-Felix serology and alter outer-membrane protein assembly. Unlike wild-type R. conorii, pso mutants cannot elicit bactericidal antibodies that bind O antigen. The pso operon is conserved among rickettsial pathogens, suggesting that bactericidal antibodies targeting O antigen may generate universal immunity that could be exploited to develop vaccines against rickettsial diseases.

Enfermedad del injerto contra el receptor tras infección por Rickettsia conorii inducida por la picadura de una garrapata / Graft-versus-host disease after an infection by Rickettsia conorii induced by a tick's bite

No disponible

Nonselective Persistence of a Rickettsia conorii Extrachromosomal Plasmid during Mammalian Infection.

Scientific analysis of the genus Rickettsia is undergoing a rapid period of change with the emergence of viable genetic tools. The development of these tools for the mutagenesis of pathogenic bacteria will permit forward genetic analysis of Rickettsia pathogenesis. Despite these advances, uncertainty still remains regarding the use of plasmids to study these bacteria in in vivo mammalian models of infection, namely, the potential for virulence changes associated with the presence of extrachromosomal DNA and nonselective persistence of plasmids in mammalian models of infection. Here, we describe the transformation of Rickettsia conorii Malish 7 with the plasmid pRam18dRGA[AmTrCh]. Transformed R. conorii stably maintains this plasmid in infected cell cultures, expresses the encoded fluorescent proteins, and exhibits growth kinetics in cell culture similar to those of nontransformed R. conorii. Using a well-established murine model of fatal Mediterranean spotted fever, we demonstrate that R. conorii(pRam18dRGA[AmTrCh]) elicits the same fatal outcomes in animals as its untransformed counterpart and, importantly, maintains the plasmid throughout infection in the absence of selective antibiotic pressure. Interestingly, plasmid-transformed R. conorii was readily observed both in endothelial cells and within circulating leukocytes. Together, our data demonstrate that the presence of an extrachromosomal DNA element in a pathogenic rickettsial species does not affect either in vitro proliferation or in vivo infectivity in models of disease and that plasmids such as pRam18dRGA[AmTrCh] are valuable tools for the further genetic manipulation of pathogenic rickettsiae.

Mediterranean spotted fever in a patient with Crohn's disease under adalimumab: First case report and review of the literature / Fiebre botonosa en un paciente con enfermedad de Crohn con tratamiento de adalimumab: Observación clínica y revisión de la literatura

No disponible

Fiebre botonosa mediterránea: a propósito de un caso / Mediterranean spotted fever: a case report

Los síntomas típicos de la fiebre botonosa mediterránea (FBM) incluyen fiebre, un rash cutáneo generalizado y la presencia de una escara negra que puede pasar desapercibida. Por lo general sigue un curso benigno, aunque ocasionalmente aparecen complicaciones graves. A pesar de que la enfermedad fue descrita hace un siglo, algunas preguntas acerca del vector y el reservorio real de la enfermedad siguen sin aclararse. Presentamos un caso típico de FBM y planteamos una hipótesis sobre la forma de transmisión de la Rickettsia en nuestro paciente

Typical symptoms of Mediterranean spotted fever (FBM) include fever, generalized rash and the presence of a black eschar that may go unnoticed. Usually follows a benign course, but occasionally serious complications occur. Although the disease was first described a century ago, a few questions about the actual vector and reservoir of the disease remain unclear. We present a typical case of FBM and propose a hypothesis about the mode of transmission of Rickettsia in our patient

Host-directed antimicrobial drugs with broad-spectrum efficacy against intracellular bacterial pathogens.

We sought a new approach to treating infections by intracellular bacteria, namely, by altering host cell functions that support their growth. We screened a library of 640 Food and Drug Administration (FDA)-approved compounds for agents that render THP-1 cells resistant to infection by four intracellular pathogens. We identified numerous drugs that are not antibiotics but were highly effective in inhibiting intracellular bacterial growth with limited toxicity to host cells. These compounds are likely to target three kinds of host functions: (i) G protein-coupled receptors, (ii) intracellular calcium signals, and (iii) membrane cholesterol distribution. The compounds that targeted G protein receptor signaling and calcium fluxes broadly inhibited Coxiella burnetii, Legionella pneumophila, Brucella abortus, and Rickettsia conorii, while those directed against cholesterol traffic strongly attenuated the intracellular growth of C. burnetii and L. pneumophila. These pathways probably support intracellular pathogen growth so that drugs that perturb them may be therapeutic candidates. Combining host- and pathogen-directed treatments is a strategy to decrease the emergence of drug-resistant intracellular bacterial pathogens. Importance: Although antibiotic treatment is often successful, it is becoming clear that alternatives to conventional pathogen-directed therapy must be developed in the face of increasing antibiotic resistance. Moreover, the costs and timing associated with the development of novel antimicrobials make repurposed FDA-approved drugs attractive host-targeted therapeutics. This paper describes a novel approach of identifying such host-targeted therapeutics against intracellular bacterial pathogens. We identified several FDA-approved drugs that inhibit the growth of intracellular bacteria, thereby implicating host intracellular pathways presumably utilized by bacteria during infection.

Compartmentalized, functional role of angiogenin during spotted fever group rickettsia-induced endothelial barrier dysfunction: evidence of possible mediation by host tRNA-derived small noncoding RNAs.

BACKGROUND: Microvascular endothelial barrier dysfunction is the central enigma in spotted fever group (SFG) rickettsioses. Angiogenin (ANG) is one of the earliest identified angiogenic factors, of which some are relevant to the phosphorylation of VE-cadherins that serve as endothelial adherens proteins. Although exogenous ANG is known to translocate into the nucleus of growing endothelial cells (ECs) where it plays a functional role, nuclear ANG is not detected in quiescent ECs. Besides its nuclear role, ANG is thought to play a cytoplasmic role, owing to its RNase activity that cleaves tRNA to produce small RNAs. Recently, such tRNA-derived RNA fragments (tRFs) have been shown to be induced under stress conditions. All these observations raise an intriguing hypothesis about a novel cytoplasmic role of ANG, which is induced upon infection with Rickettsia and generates tRFs that may play roles in SFG rickettsioses. METHODS: C3H/HeN mice were infected intravenously with a sublethal dose of R. conorii. At days 1, 3, and 5 post infection (p.i.), liver, lung and brain were collected for immunofluorescence (IF) studies of R. conorii and angiogenin (ANG). Human umbilical vein endothelial cells (HUVECs) were infected with R. conorii for 24, 48, and 72 hrs before incubation with 1µg/ml recombinant human ANG (rANG) in normal medium for 2 hrs. HUVEC samples were subjected to IF, exogenous ANG translocation, endothelial permeability, and immunoprecipitation phosphorylation assays. To identify small non-coding RNAs (sncRNAs) upon rickettsial infection, RNAs from pulverized mouse lung tissues and HUVECs were subjected to library preparation and deep sequencing analysis using an Illumina 2000 instrument. Identified sncRNAs were confirmed by Northern hybridization, and their target mRNAs were predicted in silico using BLAST and RNA hybrid programs. RESULTS: In the present study, we have demonstrated endothelial up-regulation of ANG, co-localized with SFG rickettsial infection in vivo. We also have provided direct evidence that rickettsial infection sensitizes human ECs to the translocation of exogenous ANG in a compartmentalized pattern at different times post-infection. Typically, exogenous ANG translocates into the nucleus at 24 hrs and to the cytoplasm at 72 hrs post-infection. The ANG cytoplasmic translocation enhances phosphorylation and destabilization of VE-cadherin and attenuates endothelial barrier function. Of note, deep sequencing analysis detected tRFs, mostly derived from the 5'-halves of host tRNAs, that are induced by ANG. Northern hybridization validates the two most abundantly cloned tRFs derived from tRNA-ValGTG and tRNA-GlyGCC, in both mouse tissues and human cells. Bioinformatics analysis predicted that these tRFs may interact with transcripts associated with the endothelial barrier, the host cell inflammatory response, and autophagy. CONCLUSIONS: Our data provide new insight into the role of compartmentalized ANG during SFG rickettsioses, and highlight its possible mediation through tRFs.

¿Dónde está la garrapata? Crucero & casa (resuelto en Atención Primaria) / Where is the tick? Cruise & home (resolved in primary health care)

Trabajar en Atención Primaria supone pensar en las patologías más frecuentes que se dan en nuestro entorno. Un paciente joven que presenta úlceras genitales nos hace pensar en una infección de transmisión sexual. Sin embargo, la falta de respuesta al tratamiento empírico correcto y los resultados de la serología muestran el diagnóstico definitivo: fiebre botonosa mediterránea, con una localización poco común. Debemos recordar que una enfermedad común en nuestro medio puede parecerse a otra igualmente común, por lo que una correcta historia clínica será fundamental para distinguirlas (AU)

Working in primary health care means thinking about the most common pathologies which we encounter in our workplace. A young patient with genital ulcers makes us think of a sexually transmitted infection. However, the lack of response to the correct empirical treatment and serology results show a definitive diagnosis: Mediterranean spotted fever (also known as boutonneuse fever), with a rare location. It should be remembered that a commonly occurring disease in our environment may resemble another equally common one, thus a correct clinical history is fundamental in order to distinguish them (AU)

Molecular characterization of mediterranean spotted fever rickettsia isolated from a European traveler in the state of São Paulo, Brazil.

Rickettsial spotted fever is common in southeastern Brazil. Differential diagnosis of pathogens can be performed with proper laboratory methods. A traveler arriving from Portugal developed a fatal febrile hemorrhagic syndrome diagnosed as spotted fever rickettsiosis. We isolated the agent, which was identified as Rickettsia conorii conorii by sequencing rickettsial genes.

OmpA-mediated rickettsial adherence to and invasion of human endothelial cells is dependent upon interaction with α2ß1 integrin.

Rickettsia conorii, a member of the spotted fever group (SFG) of the genus Rickettsia and causative agent of Mediterranean spotted fever, is an obligate intracellular pathogen capable of infecting various mammalian cell types. SFG rickettsiae express two major immunodominant surface cell antigen (Sca) proteins, OmpB (Sca5) and OmpA (Sca0). While OmpB-mediated entry has been characterized, the contribution of OmpA has not been well defined. Here we show OmpA expression in Escherichia coli is sufficient to mediate adherence to and invasion of non-phagocytic human endothelial cells. A recombinant soluble C-terminal OmpA protein domain (954-1735) with predicted structural homology to the Bordetella pertussis pertactin protein binds mammalian cells and perturbs R. conorii invasion by interacting with several mammalian proteins including ß1 integrin. Using functional blocking antibodies, small interfering RNA transfection, and mouse embryonic fibroblast cell lines, we illustrate the contribution of α2ß1 integrin as a mammalian ligand involved in R. conorii invasion of primary endothelial cells. We further demonstrate that OmpA-mediated attachment to mammalian cells is in part dependent on a conserved non-continuous RGD motif present in a predicted C-terminal 'pertactin' domain in OmpA.Our results demonstrate that multiple adhesin-receptor pairs are sufficient in mediating efficient bacterial invasion of R. conorii.

Genome sequence of Rickettsia conorii subsp. caspia, the agent of Astrakhan fever.

Rickettsia conorii subsp. caspia is the agent of Astrakhan fever, a spotted fever group rickettsiosis endemic to Astrakhan, Russia. The present study reports the draft genome of Rickettsia conorii subsp. caspia strain A-167.

Cross-protection among Rickettsia species and subspecies in a guinea pig model of cutaneous infection.

Pathogenic rickettsiae of the spotted fever group (SFG) induce skin lesions called "eschars" at the sites of arthropod bites. We recently described a guinea pig model based on eschar formation to predict the eventual pathogenicity of new Rickettsia species, and in this study, we used the model to study cross-protection among SFG Rickettsia species and subspecies. We showed that the intradermal inoculation of guinea pigs with Rickettsia conorii subsp. conorii significantly reduced, but not completely prevented, the number of eschars after sub-challenges with the same pathogen (P=0.0004). The same effect was also observed for other subspecies of the R. conorii complex. Additionally, the bacterial load in the eschars was significantly lower in immunized animals than in naïve animals. No protection was observed when sub-challenges were performed with other Rickettsia species, such as R. africae, R. sibirica subsp. mongolitimonae, R. aeschlimannii and R. massiliae. Our data suggest that patients may experience several episodes of infection with related or with the same species of Rickettsia. Moreover, the absence of cross-protection between Rickettsia species may explain the co-existence of two or more tick-borne rickettsioses in the same geographic areas.

Imported rickettsioses in Italy.

BACKGROUND: In Italy few cases of rickettsioses have been reported in travellers and autochthonous cases are attributed predominantly to Rickettsia conorii, the agent of Mediterranean spotted fever. METHOD: The purpose of this study was to investigate some epidemiological and clinical features of tick-borne spotted fever group rickettsiosis acquired abroad or in Italy. Serum specimens collected prospectively from patients with suspected rickettsioses were tested by immunofluorescence assay. A definitive diagnosis was made on the basis of positive serological test results at the WHO collaborative centre for rickettsial diseases, Marseille, France. We compared the clinical features of patients with confirmed rickettsioses and those showing typical clinical symptoms/signs without definitive diagnose. RESULTS: Eight of 26 patients suspected cases had confirmed rickettsioses. All patients were travellers returning from southern Africa (75% Rickettsia africae). Inoculation eschars were significantly more common in patients with confirmed rickettsioses (p = 0.004). CONCLUSIONS: Our study demonstrates that R. africae is the most frequent rickettsia observed in Italian travellers. Prior to receiving the laboratory results, physicians should start empirical treatment on the basis of epidemiologic data (e.g., travel history to Africa), and clinical findings compatible with rickettsioses (e.g., eschars).

Regulation of inducible heme oxygenase and cyclooxygenase isozymes in a mouse model of spotted fever group rickettsiosis.

Vascular endothelial cells (ECs) lining the blood vessels are the preferred primary targets of pathogenic Rickettsia species in the host. In response to oxidative stress triggered by infection, ECs launch defense mechanisms such as expression of heme oxygenase-1 (HO-1). Previous evidence from an established animal model of Rocky Mountain spotted fever also suggests selective modulation of anti-oxidant enzyme activities in the target host tissues. In this study, we have examined the expression profiles of HO-1 and COX-2 in different tissues during Rickettsia conorii infection of susceptible C3H/HeN mice. RNA hybridization with murine HO-1 and COX-2-specific complementary DNA probes revealed increased HO-1 expression in the liver and brain of mice infected with three different doses of R. conorii ranging from 2.25×10(3) to 2.25×10(5) pfu, relatively non-remarkable changes in the lungs, and a trend for down-regulation in the spleen. The most prominent HO-1 response was evident in the liver with ∼4-fold increase on day 4 post-infection, followed by a decline on day 7. HO-1 expression in the brain, however, peaked with significantly higher levels on day 7. Following infection with both sub-lethal as well as lethal doses of infection, the transcript encoding COX-2 also displayed a pattern of increased expression in the liver and brain. Although immunohistochemical staining revealed increased abundance of HO-1 protein in the liver of infected mice, adjoining serial sections did not exhibit positive staining for COX-2 in infected tissues. The levels of monocyte chemoattractant protein-1 (MCP-1) and keratinocyte-derived cytokine (KC) were significantly higher in the sera of infected mice and corresponded with the onset and severity of the disease. Treatment of infected animals with anti-oxidants α-lipoic acid and N-acetylcysteine and HO inhibitor stannous protoporphyrin (SnPPIX) showed only selective beneficial effects on HO-1 and COX-2 expression in the liver and spleen and serum levels of KC and MCP-1. R. conorii infection of susceptible mice, therefore, results in selective regulation of the expression of HO-1 and COX-2 in a manner dependent on the target host tissue's cellular environment and the propensity of infection with rickettsiae.

Deleterious effect of ciprofloxacin on Rickettsia conorii-infected cells is linked to toxin-antitoxin module up-regulation.

OBJECTIVES: To confirm and better understand the deleterious effect of fluoroquinolones reported during Rickettsia conorii infection in humans. METHODS: We used a new plaque assay to test the effect of ciprofloxacin on cells infected by R. conorii. Controls were mock-treated infected cells and infected cells treated with doxycycline. We used real-time quantitative RT-PCR to quantify vapC and vapB transcripts in cells infected by R. conorii treated with ciprofloxacin or mock treated. RESULTS: By plaque assay, at baseline (0 h) there is no difference in lytic area between cells treated with ciprofloxacin (whatever concentration used) and controls. Ciprofloxacin at 0.5 and 50 mg/L induced a significant increase in lytic areas compared with the control at 2 h, 4 h, 6 h (P<0.0001) and 24 h (P<0.0001 and P=0.035, respectively) when not induced with doxycycline. By real-time quantitative RT-PCR, ciprofloxacin was found to cause an up-regulation of toxin-antitoxin (TA) module transcription. Indeed, the mRNA levels of vapC and vapB were significantly increased at 2 h and 4 h in cells treated with 50 mg/L ciprofloxacin (not with 0.5 mg/L ciprofloxacin) compared with control levels (fold change >2.9). CONCLUSIONS: These in vitro findings correlated with our previous clinical findings: fluoroquinolones have a deleterious effect during R. conorii infection, not found with doxycycline. We speculate that the toxic effect of fluoroquinolones on R. conorii-infected cells is mediated by the overexpression of TA, possibly followed by their release into the host cytoplasm as described in Rickettsia felis.

Analysis of risk factors for malignant Mediterranean spotted fever indicates that fluoroquinolone treatment has a deleterious effect.

OBJECTIVES: To identify risk factors for malignant Mediterranean spotted fever (MSF) caused by Rickettsia conorii conorii. PATIENTS AND METHODS: Epidemiological, clinical and biological characteristics as well as risk factors (including treatment regimens) for severe MSF cases were analysed retrospectively. A patient with two or more organ dysfunctions or patient death was defined as a severe case. RESULTS: During the study period (January 1999 to December 2009), 161 MSF cases were referred to our centre for rickettsioses. Twenty-six cases (16.1%) were considered severe, which is 3-fold higher than in our previous studies. The clinical and laboratory findings were comparable to those reported elsewhere except that the type of antibiotic treatment was associated with disease severity. Doxycycline administration prior to deterioration of disease (in 31 patients) protected patients from development of severe MSF [relative risk (RR) 0.248, 95% confidence interval (CI) 0.08-0.76] and induced earlier defervescence compared with the other treatment regimens (3.02 ± 2.2 days versus 7.1 ± 6.57 days, P = 0.021). In contrast, fluoroquinolone treatment (in 21 patients) was significantly and independently associated with MSF severity (RR 2.53, 95% CI 1.40-4.55) and was associated with a significantly longer hospital stay. CONCLUSIONS: In this retrospective study fluoroquinolone treatment was associated with increased MSF disease severity. Fluoroquinolones have been previously associated with treatment failure in typhus and scrub typhus cases. Thus, we do not recommend the use of fluoroquinolones to treat rickettsial diseases.

The Rickettsia conorii autotransporter protein Sca1 promotes adherence to nonphagocytic mammalian cells.

The pathogenesis of spotted fever group (SFG) Rickettsia species, including R. conorii and R. rickettsii, is acutely dependent on adherence to and invasion of host cells, including cells of the mammalian endothelial system. Bioinformatic analyses of several rickettsia genomes revealed the presence of a cohort of genes designated sca genes that are predicted to encode proteins with homology to autotransporter proteins of Gram-negative bacteria. Previous work demonstrated that three members of this family, rOmpA (Sca0), Sca2, and rOmpB (Sca5) are involved in the interaction with mammalian cells; however, very little was known about the function of other conserved rickettsial Sca proteins. Here we demonstrate that sca1, a gene present in nearly all SFG rickettsia genomes, is actively transcribed and expressed in R. conorii cells. Alignment of Sca1 sequences from geographically diverse SFG Rickettsia species showed that there are high degrees of sequence identity and conservation of these sequences, suggesting that Sca1 may have a conserved function. Using a heterologous expression system, we demonstrated that production of R. conorii Sca1 in the Escherichia coli outer membrane is sufficient to mediate attachment to but not invasion of a panel of cultured mammalian epithelial and endothelial cells. Furthermore, preincubation of a recombinant Sca1 peptide with host cells blocked R. conorii cell association. Together, these results demonstrate that attachment to mammalian cells can be uncoupled from the entry process and that Sca1 is involved in the adherence of R. conorii to host cells.

Nen amb adenopaties doloroses occipitals i febreta: TIBOLA / A child with tender occipital limph nodes the low-grade fever: Tibola

Las enfermedades transmitidas por garrapatas son poco frecuentesen nuestro medio, aunque el antecedente de picadura y la presenciade síntomas como la fiebre y el rash obligan a descartar lafiebre botonosa o la enfermedad de Lyme.Una enfermedad emergente es la llamada TIBOLA o DEBONEL, quese presenta como una linfadenitis occipital con escara en el cuerocabelludo. Es una enfermedad transmitida por garrapatas que sepuede diagnosticar por las características clínicas que presenta.Se describe el caso de un niño de 9 años que consulta por adenopatíasdolorosas en zona occipital y cervical posterior y febrícula.Nen amb adenopaties doloroses occipitalsi febreta: TIBOLAAsunción Clopés, Rosa Rovira, Manuel Andrés Samper, Francisco J. Sanchís, Luis-EnriqueDelgado, Pilar TerradasServei de Pediatria. Pius Hospital de Valls. Valls. TarragonaEl diagnóstico de sospecha ante la presencia de escara en el cuerocabelludo y el antecedente de picadura de garrapata se confirmópor una serología cruzada positiva a Rickettsia conorii. Se efectuótratamiento con eritromicina con buena respuesta clínica y conuna alopecia cicatricial como secuela.Esta enfermedad está provocada por Rickettsia slovaca, germentransmitido por garrapatas, siendo un cuadro con pocas referenciasen nuestro medio, habitualmente por confundir la reacciónadenopática con una sobreinfección bacteriana.Ante un paciente con adenitis occipital y el antecedente de picadurapor garrapata y/o escara en el cuero cabelludo el diagnósticode TIBOLA es el más probable(AU)

Tick-borne diseases are rare in our environment; for usual differentialdiagnosis of, fever and rash in the presence of history oftick-bite, boutoneusse fever or Lyme disease should be considered.TIBOLA is an emergent tick-borne disease that presents with occipitallymphadenitis and a scalp eschar; this entity should be easyto diagnose based on its clinical presentation.We describe the case of a 9 year-old boy who presented with tenderoccipital lymphadenopathies and low-grade fever. In the presenceof a scalp eschar and the history of tick-bite, the diagnosisof TIBOLA was suspected, which was confirmed with cross-positiveserology to Rickettsia conorii. Treatment with erythromycin wasadministered, with prompt improvement of the symptoms and theonly sequel of residual alopecia at the eschar site.TIBOLA is caused by Rickettsia slovaca; there are few reports ofthis entity in our region, probably because it is often misdiagnosedas bacterial lymphadenitis.TIBOLA should be suspected in any patient that presents with occipitallymphadenitis and history of tick-bite or scalp eschar(AU)